Laboratory of Neurobiology & Molecular Pharmacology, Centre de Psychiatrie et Neurosciences, UMR-894 INSERM/Université Paris Descartes, 2Ter rue d'Alésia, Paris, France.
Br J Pharmacol. 2011 Feb;162(4):947-60. doi: 10.1111/j.1476-5381.2010.01094.x.
By interacting with trkB receptors, brain-derived neurotrophic factor (BDNF) triggers various signalling pathways responsible for neurone survival, differentiation and modulation of synaptic transmission. Numerous reports have implicated BDNF and trkB in the pathogenesis of various central nervous system affections and in cancer, thus representing trkB as a promising therapeutic target. In this study, we used an antibody-based approach to search for trkB-selective functional reagents.
Six commercially available polyclonal and monoclonal antibodies were tested on recombinant and native, human and rodent trkB receptors. Functional and pharmacological characterization was performed using a modified version of the KIRA-elisa method and radioligand binding studies. Western blot analyses and neurite outgrowth assays were carried out to determine the specificity and selectivity of antibody effects. The survival properties of one antibody were further assessed on cultured neurones in a serum-deprived paradigm.
The functional trkB-selective antibodies showed distinct pharmacological profiles, ranging from partial agonists to antagonists, acting on trkB receptors through allosteric modulations. The same diversity of effects was observed on the mitogen-activated protein kinase signalling pathway downstream of trkB and on the subsequent neurite outgrowth. One antibody with partial agonist activity demonstrated cell survival properties by activating the Akt pathway. Finally, these antibodies were functionally validated as true trkB-selective ligands because they failed activating trkA or trkC, and contrary to BDNF, none of them bind to p75(NTR).
These trkB-selective antibodies represent a novel class of pharmacological tools to explore the pathophysiological roles of trkB and its potential therapeutic relevance for the treatment of various disorders.
脑源性神经营养因子（BDNF）通过与 trkB 受体相互作用，触发各种信号通路，负责神经元的存活、分化和突触传递的调节。大量报道表明 BDNF 和 trkB 在各种中枢神经系统疾病和癌症的发病机制中起作用，因此 trkB 被认为是一个有前途的治疗靶点。在这项研究中，我们使用基于抗体的方法来寻找 trkB 选择性功能试剂。
六种市售的多克隆和单克隆抗体在重组和天然的、人和啮齿动物的 trkB 受体上进行了测试。使用 KIRA-elisa 方法的修改版本和放射性配体结合研究进行功能和药理学特征分析。进行 Western blot 分析和神经突生长测定，以确定抗体作用的特异性和选择性。一种抗体的生存特性在血清剥夺范式下进一步在培养的神经元上进行评估。
功能上选择性的 trkB 抗体显示出不同的药理学特征，从部分激动剂到拮抗剂，通过变构调节作用于 trkB 受体。在 trkB 下游的丝裂原活化蛋白激酶信号通路以及随后的神经突生长方面也观察到相同的多种效应。一种具有部分激动剂活性的抗体通过激活 Akt 通路显示出细胞生存特性。最后，这些抗体被功能上验证为真正的 trkB 选择性配体，因为它们不能激活 trkA 或 trkC，而且与 BDNF 不同，它们都不与 p75（NTR）结合。
这些 trkB 选择性抗体代表了一类新的药理学工具，可以探索 trkB 的病理生理作用及其在治疗各种疾病中的潜在治疗相关性。
